Use of growth hormone fragments

ABSTRACT

The invention provides a method for treating a condition in which growth hormone administration is beneficial, for treating osteoarthritis, for increasing chondrocyte, proteoglycan or collagen production or quality or repair or promoting cartilage tissue formation or repair, for promoting or improving muscle, ligament or tendon mass, repair, form or function, or for treating inflammatory, traumatic or genetic diseases of muscle or connective tissue, comprising administering to a subject an effective amount of a peptide comprising a carboxyl-terminal sequence from a growth hormone and not the IGF-1 domain of growth hormone. It also provides a method of treating a condition involving insufficient functional chondrocytes or insufficient functional cartilage tissue, the method comprising administering to a subject in need thereof a peptide comprising a carboxyl-terminal sequence from a growth hormone and not the domain of growth hormone responsible for IGF-1 production.

FIELD

The invention relates to the use of peptide fragments of growth hormoneto treat conditions treatable by growth hormone without the side effectsassociated with use of growth hormone.

BACKGROUND

All references, including any patents or patent application, cited inthis specification are hereby incorporated by reference to enable fullunderstanding of the invention. Nevertheless, such references are not tobe read as constituting an admission that any of these documents formspart of the common general knowledge in the art, in Australia or in anyother country. The discussion of the references states what theirauthors assert, and the applicants reserve the right to challenge theaccuracy and pertinency of the cited documents.

Somatotropin or growth hormone (GH) is a potent anabolic hormoneproduced by the pituitary gland in daily doses of 0.5 to 0.8 mg inchildren and young adults. Its production decreases rapidly with age.

Human growth hormone (hGH) has a number of metabolic effects, the mostprominent of which is its anabolic effect. HGH increases the influx ofamino acids into the cell and decreases the efflux. Cell proliferationis accentuated as is overall protein synthesis and new tissue growth.HGH also stimulates insulin-like growth factor-1 (IGF-1) production bythe liver and the anabolism seen with hGH is due to the action of IGF-1.Additionally, hGH, due to IGF-1 activity, is known to accelerate nucleicacid translation and transcription, increase nitrogen retention,increase protein synthesis, decrease cortisol receptor activity,increase hydrolysis of fats to fatty acids, increase fat oxidation forfuel thereby decreasing fat stores, increase metabolic rate, causeinitial fluid retention, produce insulin resistance often leading tohyperglycemia and increase insulin requirements.

The predominant form of hGH is a globular protein with a molecularweight of 22 kilodaltons, consisting of 191 amino acid residues in asingle-chain, folded by 2 disulphide bonds with a small loop at thecarboxyl terminus between residues 182 and 189. Crystallographic studiesshow that hGH contains four anti-parallel α-helices which are arrangedin a left-twisted, tightly-packed helical bundle. The concept that thereare discrete functional domains within the hGH molecule responsible forspecific metabolic actions of the hormone is generally accepted. Theamino-terminus has been identified as the functional domain responsiblefor IGF-1 secretion and therefore the insulin-like actions of hGH (Ng(1990) New Antidiabetic Drugs pp 197-205). The carboxyl-terminus hasbeen identified as a lipolytic domain of hGH (Ng F M et al., (2000) J.Mol. Endocrin. 25, 287-298).

HGH serves as a critical hormone in the regulation of cell and organgrowth and in physiological function upon various stages of aging. Forexample, over-production of hGH results in gigantism in children andacromegaly in adults, whereas under-production leads to dwarfism inchildren, and chronic renal insufficiency. In adults, hGH deficiency canaffect metabolic processing of proteins, carbohydrates, lipids, mineralsand connective tissue and can result in muscle, bone or skin atrophy.Other hGH deficiency disorders characterized by growth failure includePrader-Willi syndrome, intrauterine growth retardation and catabolicstate for example during chemotherapy treatment and in the treatment ofAIDS.

Many scientific studies confirm that growth hormone treatment in adultsimproves the body composition (increasing the muscle mass and decreasingfat), bone density, muscle strength, cardiovascular parameters (i.e.decrease of LDL cholesterol), and the quality of life.

Advanced acquired immunodeficiency syndrome (AIDS) is often accompaniedby muscle wasting (“AIDS wasting”) and hGH has been shown to amelioratethis condition. HGH has been given to promote healing of large burns byreducing the amount of protein breakdown during the early post-injuryperiod. HGH has been used as an adjunct to caloric restriction forobesity as hGH promotes lipolysis and reduces proteolysis. Severalstudies have demonstrated improvements in exercise capacity and cardiacfunction among hGH-deficient patients receiving hGH replacement. Suchpatients show increased oxygen uptake and power output during cycleergometry associated with increased skeletal muscle mass and improvedcardiac function.

It is widely accepted that the anabolic effects of hGH are mediated viasecretion of IGF-1. Elevated IGF-1 may lead to reduced insulinsensitivity, increased hyperglycemic episodes, fluid retention,diabetes, acromegaly and some cancers. Accordingly, clinicalapplications of hGH are currently restricted.

It is an aim of an embodiment of the present invention to providetreatments for conditions that can be treated by growth hormone, withoutelevation of IGF-1.

SUMMARY

A first aspect provides a method for:

-   -   (a) treating a condition in which growth hormone administration        is beneficial,    -   (b) treating osteoarthritis,    -   (c) increasing chondrocyte, proteoglycan or collagen production        or quality or repair or promoting cartilage tissue formation or        repair,    -   (d) promoting or improving muscle, ligament or tendon mass,        repair, form or function, or    -   (e) treating inflammatory, traumatic or genetic diseases of        muscle or connective tissue, comprising administering to a        subject an effective amount of a peptide comprising a        carboxyl-terminal sequence from a growth hormone and not the        domain of growth hormone responsible for IGF-1 production.

An alternative form of the first aspect provides a composition fortreatment of a condition in which growth hormone administration isbeneficial, for treating osteoarthritis, for increasing chondrocyte,proteoglycan or collagen production or quality or repair or promotingcartilage tissue formation or repair, for promoting or improving muscle,ligament or tendon mass, repair, form or function, or for treatinginflammatory, traumatic or genetic diseases of muscle or connectivetissue, the composition comprising a peptide comprising acarboxyl-terminal sequence from a growth hormone and not the domain ofgrowth hormone responsible for IGF-1 production.

A further alternative form of the first aspect provides use of a peptidecomprising a carboxyl-terminal sequence from a growth hormone and notthe domain of growth hormone responsible for IGF-1 production in themanufacture of a medicament for treating a condition in which growthhormone administration is beneficial, for treating osteoarthritis, forincreasing chondrocyte, proteoglycan or collagen production or qualityor repair or promoting cartilage tissue formation or repair, forpromoting or improving muscle, ligament or tendon mass, repair, form orfunction or for treating inflammatory, traumatic or genetic diseases ofmuscle or connective tissue.

In an embodiment of the first aspect the condition is not obesity or abone disorder.

In an embodiment of the first aspect the peptide is capable of enhancingdifferentiation into myoblasts.

A second aspect provides a method of treating a condition involvinginsufficient functional chondrocytes or insufficient functionalcartilage tissue, the method comprising administering to a subject inneed thereof a peptide comprising a carboxyl-terminal sequence from agrowth hormone and not the domain of a growth hormone responsible forIGF-1 production.

An alternative form of the second aspect provides a composition fortreating a condition involving insufficient functional chondrocytes orinsufficient functional cartilage tissue, the composition comprising aneffective amount of a peptide comprising a carboxyl-terminal sequencefrom a growth hormone, and not the domain of a growth hormoneresponsible for IGF-1 production.

A further alternative form of the second aspect provides use of apeptide comprising a carboxyl-terminal sequence from a growth hormoneand not the domain of a growth hormone responsible for IGF-1 productionin the manufacture of a medicament for treating a condition involvinginsufficient functional chondrocytes or insufficient functionalcartilage tissue.

In an embodiment of the second aspect the condition is osteoarthritis.

In an embodiment of the first and second aspects the peptide comprises acarboxyl-terminal sequence from human growth hormone or a carboxylterminal sequence from a growth hormone of a non-human animal.

In an embodiment of the first and second aspects the peptide is capableof promoting chondrocyte production or regeneration. In an embodiment ofthe first and second aspects the peptide is capable of promotingcartilage tissue production or regeneration. In an embodiment of thefirst and second aspects the peptide is capable of enhancingdifferentiation of mesenchymal cells to chondrocytes or cartilagetissue. The methods provide enhanced chondrocyte production or repair.They also enhance cartilage production or repair.

In an embodiment of the first and second aspects the peptide is capableof promoting proteoglycan production. In an embodiment of the first andsecond aspects the peptide is capable of promoting collagen production.In an embodiment of the first and second aspects the methods promoteproduction of cartilage, chondrocytes and or proteoglycans in damagedcartilage.

In an embodiment of the first and second aspects, the peptide isadministered on or in an agent, implant, medical device, prosthesis orcellular scaffold.

In an embodiment of the first and second aspects the condition is one inwhich a protein anabolic effect is beneficial, including traumarecovery, for treating burns or for promoting growth, repair, strength,form or function of muscle, tendons and ligaments.

In an embodiment of the first and second aspects, the peptide comprisesAOD9604 (SEQ ID NO: 1).

In an embodiment of the first and second aspects the peptide consistsessentially of AOD9604 (SEQ ID NO: 1).

In an embodiment of the first and second aspects administration of thepeptide or medicament does not elevate IGF-1 in serum and hence thetreatment provides therapeutic effect in the absence of side effectsassociated with elevated IGF-1 in serum.

BRIEF DESCRIPTION OF THE FIGURES

FIG. 1 shows DNA content for chondrocyte culture for 3 weeks on filterwith treatment with AOD (SEQ ID NO:1) showing treatment does not have atoxic effect on cells.

FIG. 2 shows proteoglycan content for chondrocyte culture for 3 weeks onfilter with treatment with AOD (SEQ ID NO:1).

FIG. 3 shows collagen content for chondrocyte culture for 3 weeks onfilter with treatment with AOD (SEQ ID NO:1).

FIG. 4 shows that AOD (SEQ ID NO:1) has no effect on the DNA content ofnative cartilage.

FIGS. 5 and 7 show that AOD (SEQ ID NO:1) does not affect proteoglycancontent of native cartilage.

FIGS. 6 and 8 show that AOD (SEQ ID NO:1) does not affect collagencontent of native cartilage.

FIG. 9 shows that AOD (SEQ ID NO:1) induces differentiation of C2C12cells into myoblasts after 24 hours of treatment.

FIG. 10 shows that AOD (SEQ ID NO:1) induces differentiation of C2C12cells into myoblasts after 3 days of treatment.

FIGS. 11 and 12 show that AOD (SEQ ID NO:1) treatment does not enhancemyotube Formation or alter cell proliferation.

FIG. 13 shows histological analysis of lateral and medial condyles ofthe right knee joint of a rabbit osteoarthritis model treated with Group1 (4 weekly injections of vehicle).

FIG. 14 shows histological analysis of lateral and medial condyles ofthe right knee joint of a rabbit osteoarthritis model treated with Group2 (4 weekly injections of 6 mg of HA-hyuran-plus).

FIG. 15 shows histological analysis of lateral and medial condyles ofthe right knee joint of a rabbit osteoarthritis model treated with Group3 (4 weekly injections of 0.25 mg of AOD9604).

FIG. 16 shows histological analysis of lateral and medial condyles ofthe right knee joint of a rabbit osteoarthritis model treated with Group4 (4 weekly injections of both HA and AOD9604).

DESCRIPTION

Ng, F M et al., (1990) supra identifies the amino-terminus of humangrowth hormone as the functional domain responsible for IGF-1 secretionand therefore the insulin-like actions of hGH.

In Australian patent No. 693478 by Monash University, the applicantdescribes the use of a peptide derived from the carboxyl-terminalsequence of human growth hormone, or a corresponding region from growthhormone of other mammalian species, for the control of obesity. Thisregion of growth hormone has the ability to modulate lipid metabolism.In particular, a synthetic peptide corresponding to amino acid residues177-191 of human growth hormone sequence (hereinafter referred to as hGH177-191) was found to reduce body weight gain and adipose tissue mass ina model system for obesity, the C57Bl/6J (Ob/Ob) mouse. A subsequentapplication, PCT/AU98/00724 by Metabolic Pharmaceuticals Ltd, disclosesanalogues of the hGH177-191 peptide which share this activity.PCT/AU00/01362 (WO01/33977), also by Metabolic Pharmaceuticals Limited,discloses the surprising oral activity of such peptides.

Clinical trials on a particular hGH 177-191 analogue, designated AOD9604and having the sequence provided as SEQ ID NO: 1 below, showed that theeffective dose of the peptide administered did not elevate the plasmalevel of IGF-1. Hence AOD9604 and other C-terminal growth hormonefragments were shown to have their effect on lipid metabolism distinctfrom any effect on IGF-1 secretion. This was particularly surprising asit had long been held in the art that the effects of hGH are modulatedby IGF-1 secretion. SEQ ID NO: 1Tyr-Leu-Arg-Ile-Val-Gln-Cys-Arg-Ser-Val-Glu-Gly-Ser-Cys-Gly-Phe (AOD9604or Tyr-hGH 177-191)

Further work on AOD9604 and other C-terminal growth hormone fragmentsindicated an effect on bone metabolism, particularly increasedosteoblast formation or without an increase in the formation ofosteoclasts. This was particularly unexpected as it had been long heldin the art that the effect of full length human growth hormone on bonemetabolism was through increased IGF-1 secretion, which does not occurwith administration of AOD9604. This invention is described inPCT/AU05/00638 to Metabolic Pharmaceuticals Ltd.

The entire disclosures of AUG693478, PCT/AU98/00724, PCT/AU00/01362 andPCT/AU05/00638 are incorporated herein by this reference.

The inventors have now determined that unexpectedly their C-terminalgrowth hormone fragments, particularly AOD9604 (SEQ ID NO:1) have ananabolic effect on chondrocytes and muscle. Previously it was consideredthat the anabolic effect of hGH on chondrocytes and muscle was regulatedby IGF-1 secretion by human growth hormone and that since the IGF domainwas not present in the C-terminal growth hormone fragments that Cterminal growth hormone peptides would not have an anabolic effect asthey lacked the functional domain of hGH. The inventors have shown thisis not the case, thus allowing the treatments with a C terminal growthhormone peptide to proceed without increased IGF-1 secretion and theside effects associated therewith.

For example, the inventors have now determined that C-terminal growthhormone fragments, particularly AOD9604 (SEQ ID NO:1) are capable ofincreasing chondrocyte, proteoglycan, collagen and production orquality, repair or regeneration and accordingly may improve cartilageproduction or quality. While previously it was known that human growthhormone stimulates chrondrocyte proliferation it was considered thatthese treatments resulted in increased IGF-1 secretion. Additionally theinventors have now shown that C-terminal growth hormone fragments caninduce differentiation into myoblasts therefore suggesting use of thefragments in conditions requiring muscle growth. The inventors havepreviously shown that their C-terminal growth hormone fragment does notelevate IGF plasma levels. The new findings provide therapies to proceedwithout increased IGF-1 secretion and the side effects associatedtherewith.

Side effects to be avoided include at least one of increased insulinresistance, increased insulin demand, fluid retention, hypercalcaemiaand acromegaly.

Treatments

The invention in one aspect relates to the treatment of conditions. Theterms “treating” and “treatment” as used herein refer to reduction inseverity and/or frequency of symptoms, elimination of symptoms and/orunderlying cause, prevention of the occurrence of symptoms (prophylaxis)and/or their underlying cause, and improvement or remediation of damage.Thus, for example, the present method of “treating” a conditionencompasses both prevention of the condition in a predisposedindividual, treatment of the condition in a clinically symptomaticindividual and treatment of a healthy individual for beneficial effect.

“Treating” as used herein covers any treatment of, repair or preventionof a condition in a vertebrate, a mammal, particularly a human ordomestic, bloodstock, farm or zoo animal, and includes inhibiting thecondition, i.e., arresting its development; or relieving or amelioratingthe effects of the condition, i.e., cause regression of the effects ofthe condition and producing a beneficial effect.

“Prophylaxis” or “prophylactic” or “preventative” therapy as used hereinincludes preventing the condition from occurring or ameliorating thesubsequent progression of the condition in a subject that may bepredisposed to the condition, but has not yet been diagnosed as havingit.

As used herein, “condition” refers to any deviation from normal healthand includes a disease, disorder, defect or injury, such as injurycaused by trauma, and deterioration due to age, inflammatory, infectiousor genetic disorder or due to environment.

Conditions in which growth factor administration is beneficial aredisclosed in the prior art, for example in US2011/0112021 andWO2011/038205. Of those, conditions that may be treated in accordancewith the present invention fall generally into the categories of thosein which increased chondrocyte, collagen, proteoglycan, cartilage ormuscle mass form or function is desirable. Conditions treatable by thepresent invention include osteoarthritis; rheumatoid arthritis, juvenilerheumatoid arthritis; HIV-infection in children receiving HAARTtreatment (HIV/HALS children); skeletal dysplasia; hypochondroplasia;achondroplasia; treatment of patients after tendon or ligament surgeryin hand, knee, or shoulder; distraction osteogenesis; disordersresulting from hip or discus replacement, meniscus repair, spinalfusions or prosthesis fixation, such as in the knee, hip, shoulder,elbow, wrist or jaw; disorders resulting from fixing of osteosynthesismaterial, such as nails, screws and plates; non-union or mal-union offractures; disorders resulting from osteatomia, e.g. from tibia or 1sttoe; disorders resulting from graft implantation; articular cartilagedegeneration in knee caused by trauma or arthritis; adult patients inchronic dialysis (APCD); malnutritional associated cardiovasculardisease in APCD; reversal of cachexia in APCD; chronic abstractivepulmonal disease in APCD; HIV in APCD; elderly with APCD; chronic liverdisease in APCD, fatigue syndrome in APCD; impaired liver function;males with HIV infections; HIV-associated lipodystrophy syndrome (HALS);male infertility; treatment of patients after major elective surgery,alcohol/drug detoxification or neurological trauma; aging; frailelderly; traumatically damaged cartilage; fibromyalgia; memorydisorders; traumatic brain injury; subarachnoid haemorrhage; very lowbirth weight; myocardial fibrosis; idiopathic dilated cardiomyopathy; aneurological disease or disorder; neuro-degeneration; delayed orimpaired growth; a cardiovascular disease or disorder; a viral syndrome(such as AIDS); diabetes; complications of diabetes (e.g., retinopathy);anorexia; bulimia; cancer cachexia; AIDS; AIDS wasting; cachexia;wasting; a renal disease or disorder; an inflammatory disease ordisorder; inflammation; Prader-Willi syndrome (PWS); chronic renalinsufficiency (CRI); aging; end-stage Renal Failure; End stage renaldisease (ESRD); Cystic Fibrosis; Erectile dysfunction; HIVlipodystrophy; skeletal dysplasia; Noonan's syndrome; glucocorticoidmyopathy; infection; diabetes; hypertension; multiple sclerosis; heartfailure; hematoma, ulcerative colitis and burns.

As used therein “neurological disease or disorder” refers to any diseaseor disorder of the nervous system and or visual system exceptdepression. “Neurological disease or disorder” include disease ordisorders that involve the central nervous system (brain, brainstem andcerebellum), the peripheral nervous system (including cranial nerves),and the autonomic nervous system (parts of which are located in bothcentral and peripheral nervous system). Examples of neurologicaldisorders include but are not limited to, headache, stupor and coma,dementia, seizure, sleep disorders, trauma, infections, neoplasms,neuro-opthalmology, movement disorders, demyelinating diseases, spinalcord disorders, and disorders of peripheral nerves, muscle andneuromuscular junctions. The following is a list of several neurologicaldisorders, symptoms, signs and syndromes that may be treated accordingto the present invention: acquired epileptiform aphasia; acutedisseminated encephalomyelitis; adrenoleukodystrophy; age-relatedmacular degeneration; agenesis of the corpus callosum; agnosia; Aicardisyndrome; Alexander disease; Alpers1 disease; alternating hemiplegia;Vascular dementia; amyotrophic lateral sclerosis; anencephaly; Angelmansyndrome; angiomatosis; anoxia; aphasia; apraxia; arachnoid cysts;arachnoiditis; AnronI-Chiari malformation; arteriovenous malformation;Asperger syndrome; ataxia telegiectasia; attention deficit hyperactivitydisorder, autism; autonomic dysfunction; back pain; Batten disease;Behcet's disease; Bell's palsy; benign essential blepharospasm; benignfocal; amyotrophy; benign intracranial hypertension; Binswanger'sdisease; blepharospasm; Bloch Sulzberger syndrome; brachial plexusinjury; brain abscess; brain injury; brain tumors (includingglioblastoma multiforme); spinal tumor; Brown-Sequard syndrome; Canavandisease; carpal tunnel syndrome; causalgia; central pain syndrome;central pontine myelinolysis; cephalic disorder; cerebral aneurysm;cerebral arteriosclerosis; cerebral atrophy; cerebral gigantism;cerebral palsy; Charcot-Marie-Tooth disease; chemotherapy-inducedneuropathy and neuropathic pain; chorea; chronic inflammatorydemyelinating polyneuropathy; chronic pain; chronic regional painsyndrome; Coffin Lowry syndrome; coma, including persistent vegetativestate; congenital facial diplegia; corticobasal degeneration; cranialarteritis; craniosynostosis; Creutzfeldt-Jakob disease; cumulativetrauma disorders; Cushing's syndrome; cytomegalic inclusion bodydisease; cytomegalovirus infection; dancing eyes-dancing feet syndrome;DandyWalker syndrome; Dawson disease; De Morsier's syndrome;Dejerine-Klumke palsy; dementia; dermatomyositis; diabetic neuropathy;diffuse sclerosis; dysautonomia; dysgraphia; dyslexia; dystonias; earlyinfantile epileptic encephalopathy; empty sella syndrome; encephalitis;encephaloceles; encephalotrigeminal angiomatosis; epilepsy; Erb's palsy;essential tremor; Fabry's disease; Fahrs syndrome; fainting; familialspastic paralysis; febrile seizures; Fisher syndrome; Friedreich'sataxia; fronto-temporal dementia and other “tauopathies”; Gaucherdisease; Gerstmann's syndrome; giant cell arteritis; giant cellinclusion disease; globoid cell leukodystrophy; Guillain-Barre syndrome;HTLV-1-associated myelopathy; Hallervorden-Spatz disease; head injury;headache; hemifacial spasm; hereditary spastic paraplegia; heredopathiaatactic a polyncuriuformis; herpes zoster oticus; herpes zoster;Hirayama syndrome; HIV-associated dementia and neuropathy;holoprosencephaly; Huntington's disease and other polyglutamine repeatdiseases; hydranencephaly; hydrocephalus; hypercortisolism; hypoxia;immune-mediated encephalomyelitis; inclusion body myositis;incontinentia pigmenti; infantile phytanic acid storage disease;infantile refsum disease; infantile spasms; inflammatory myopathy;intracranial cyst; intracranial hypertension; Joubert syndrome;Keams-Sayre syndrome; Kennedy disease Kinsboume syndrome; Klippel Feilsyndrome; Krabbe disease; Kugelberg-Welander disease; kuru; Laforadisease; Lambert-Eaton myasthenic syndrome; Landau-Kleflher syndrome;lateral medullary (Wallenberg) syndrome; learning disabilities; Leigh'sdisease; Lennox-Gustaut syndrome; Lesch-Nyhan syndrome; leukodystrophy;Lewy body dementia; Lissencephaly; locked-in syndrome; Lou Gehrig'sdisease (i.e., motor neuron disease or amyotrophic lateral sclerosis);lumbar disc disease; Lyme disease—neurological sequelae; Machado-Josephdisease; macrencephaly; megalencephaly; Melkersson-Rosenthal syndrome;Menieres disease; meningitis; Menkes disease; metachromaticleukodystrophy; microcephaly; migraine; Miller Fisher syndrome;mini-strokes; mitochondrial myopathies; Mobius syndrome; monomelicamyotrophy; motor neuron disease; Moyamoya disease;mucopolysaccharidoses; milti-infarct dementia; multifocal motorneuropathy; multiple sclerosis and other demyelinating disorders;multiple system atrophy with postural hypotension; muscular dystrophy;myasthenia gravis; myelinoclastic diffuse sclerosis; myoclonicencephalopathy of infants; myoclonus; myopathy; myotonia congenital;narcolepsy; neurofibromatosis; neuroleptic malignant syndrome;neurological manifestations of AIDS; neurological sequelae oflupus;neuromyotonia; neuronal ceroid lipofuscinosis; neuronal migrationdisorders; Niemann-Pick disease; O'Sullivan-McLeod syndrome; occipitalneuralgia; occult spinal dysraphism sequence; Ohtahara syndrome;olivopontocerebellar atrophy; opsoclonus myoclonus; optic neuritis;orthostatic hypotension; overuse syndrome; paresthesia;Neurodegenerative disease or disorder (Parkinson's disease, Huntington'sdisease, Alzheimer's disease, amyotrophic lateral sclerosis (ALS),dementia, multiple sclerosis and other diseases and disorders associatedwith neuronal cell death); paramyotonia congenital; paraneoplasticdiseases; paroxysmal attacks; Parry Romberg syndrome;Pelizaeus-Merzbacher disease; periodic paralyses; peripheral neuropathy;painful neuropathy and neuropathic pain; persistent vegetative state;pervasive developmental disorders; photic sneeze reflex; phytanic acidstorage disease; Pick's disease; pinched nerve; pituitary tumors;polymyositis; porencephaly; post-polio syndrome; postherpetic neuralgia;postinfectious encephalomyelitis; postural hypotension; Prader-Willisyndrome; primary lateral sclerosis; prion diseases; progressivehemifacial atrophy; progressive multifocalleukoencephalopathy;progressive sclerosing poliodystrophy; progressive supranuclear palsy;pseudotumor cerebri; Ramsay-Hunt syndrome (types I and 11); Rasmussen'sencephalitis; reflex sympathetic dystrophy syndrome; Refsum disease;repetitive motion disorders; repetitive stress injuries; restless legssyndrome; retrovirus-associated myelopathy; Rett syndrome; Reye'ssyndrome; Saint Vitus dance; Sandhoff disease; Schilder's disease;schizencephaly; septo-optic dysplasia; shaken baby syndrome; shingles;Shy-Drager syndrome; Sjogren's syndrome; sleep apnea; Soto's syndrome;spasticity; spina bifida; spinal cord injury; spinal cord tumors; spinalmuscular atrophy; Stiff-Person syndrome; stroke; Sturgc-Wcbcr syndrome;subacute sclerosing panencephalitis; subcortical arterioscleroticencephalopathy; Sydenham chorea; syncope; syringomyelia; tardivedyskinesia; Tay-Sachs disease; temporal arteritis; tethered spinal cordsyndrome; Thomsen disease; thoracic outlet syndrome; Tic Douloureux;Todd's paralysis; Tourette syndrome; transient ischemic attack;transmissible spongiform encephalopathies; transverse myelitis;traumatic brain injury; tremor; trigeminal neuralgia; tropical spasticparaparesis; tuberous sclerosis; vascular dementia (multi-infarctdementia); vasculitis including temporal arteritis; Von Hippel-Lindaudisease; Wallenberg's syndrome; Wera“nig-Hoffman disease; West syndrome;whiplash; Williams syndrome; Wildon's disease; and Zellweger syndrome.

As used therein an “inflammation” refers to systemic inflammatoryconditions and conditions associated locally with migration andattraction of monocytes, leukocytes and/or neutrophils. Examples ofinflammation include, but are not limited to, inflammation resultingfrom infection with pathogenic organisms (including gram-positivebacteria, gram-negative bacteria, viruses, fungi, and parasites such asprotozoa and helminths), transplant rejection (including rejection ofsolid organs such as kidney, liver, heart, lung or cornea, as well asrejection of bone marrow transplants including graft-versus-host disease(GVHD)), or from localized chronic or acute autoimmune disease orallergic reactions.

Autoimmune diseases include acute glomerulonephritis; rheumatoid orreactive arthritis; chronic glomerulonephritis; inflammatory boweldiseases such as Crohn's disease, ulcerative colitis and necrotizingenterocolitis; hepatitis; sepsis; alcoholic liver disease; non-alcoholicsteatosis; granulocyte transfusion associated syndromes; inflammatorydermatoses such as contact dermatitis, atopic dermatitis, psoriasis;systemic lupus erythematosus (SLE), autoimmune thyroiditis, multiplesclerosis, and some forms of diabetes, or any other autoimmune statewhere attack by the subject's own immune system results in pathologictissue destruction.

Allergic reactions include allergic asthma, chronic bronchitis, acuteand delayed hypersensitivity.

Systemic inflammatory disease states include inflammation associatedwith trauma, burns, reperfusion following ischemic events (e.g.thrombotic events in heart, brain, intestines or peripheral vasculature,including myocardial infarction and stroke), sepsis, ARDS or multipleorgan dysfunction syndrome. Inflammatory cell recruitment also occurs inatherosclerotic plaques.

Inflammation includes, but is not limited to, Non-Hodgkin's lymphoma,Wegener's granulomatosis, Hashimoto's thyroiditis, hepatocellularcarcinoma, thymus atrophy, chronic pancreatitis, rheumatoid arthritis,reactive lymphoid hyperplasia, osteoarthritis, ulcerative colitis,papillary carcinoma, Crohn's disease, ulcerative colitis, acutecholecystitis, chronic cholecystitis, cirrhosis, chronic sialadenitis,peritonitis, acute pancreatitis, chronic pancreatitis, chronicGastritis, adenomyosis, endometriosis, acute cervicitis, chroniccervicitis, lymphoid hyperplasia, multiple sclerosis, hypertrophysecondary to idiopathic thrombocytopenic purpura, primary IgAnephropathy, systemic lupus erythematosus, psoriasis, pulmonaryemphysema, chronic pyelonephritis, and chronic cystitis. Acardiovascular disease or disorder includes those disorders that caneither cause ischemia or are caused by reperfusion of the heart.Examples include, but are not limited to, atherosclerosis, coronaryartery disease, granulomatous myocarditis, chronic myocarditis(non-granulomatous), primary hypertrophic cardiomyopathy, peripheralartery disease (PAD), peripheral vascular disease, venousthromboembolism, pulmonary embolism, stroke, angina pectoris, myocardialinfarction, cardiovascular tissue damage caused by cardiac arrest,cardiovascular tissue damage caused by cardiac bypass, cardiogenicshock, and related conditions that would be known by those of ordinaryskill in the art or which involve dysfunction of or tissue damage to theheart or vasculature, especially, but not limited to, tissue damagerelated to GH activation.

Cardiovascular diseases include, but are not limited to,atherosclerosis, granulomatous myocarditis, myocardial infarction,myocardial fibrosis secondary to valvular heart disease, myocardialfibrosis without infarction, primary hypertrophic cardiomyopathy, andchronic myocarditis (non-granulomatous).

Other treatments proposed for growth hormone and thus the presentinvention include nerve repair, pain relief, nerve repair and nervegrowth. Prior art also proposes that growth hormone induces keratingrowth and thus the present invention can be used to enhance the growthof fingernails and hair and thus be a treatment for baldness andalopecia.

Persons skilled in the art will appreciate that persons affected withone or more of the conditions outlined above could have been treatedwith hGH in accordance with prior art teaching but that such treatmentwould have undesirable side effects caused by elevated IGF-1. Theinventors accordingly propose that such conditions could be treated witha C-terminal growth hormone fragment as defined herein without elevatedIGF-1 and the side effects this causes.

Additionally the inventors have shown particular uses for a C terminalgrowth hormone fragment that may or may not have been proposed for fulllength growth hormone.

Chondrocytes are the only cells found in cartilage. They produce andmaintain the cartilaginous matrix, which consists mainly of Type IIcollagen, proteoglycans and elastin.

Cartilage is a flexible connective tissue found in many areas in thebodies of humans and animals, including joints between bones, rib cage,ear, nose, elbow, knee, ankle, bronchial tubes and intervertebral discs.Unlike other connective tissues, cartilage does not contain bloodvessels and thus has limited repair capabilities. Because chondrocytesare bound in lacunae, they cannot migrate to damaged areas. Therefore,if cartilage is damaged, it is difficult and slow to heal.

For the purpose of the present disclosure, conditions that can betreated include Chondrocyte-Related Conditions that will benefit fromrepair or new growth of cartilage tissue or chondrocytes. This is notexclusive however and is used descriptively to emphasise the benefit ofthe presently disclosed methods.

Chondrocyte-Related Conditions include joint disorders involvingcartilage damage and include cartilage damage caused by tibial plateaudecompression.

The cause of osteoarthritis is multifactorial and includes body habitus,genetics and hormonal status.

In osteoarthritis, the cartilage covering bones (articular cartilage—asubset of hyaline cartilage) is thinned, eventually completely wearingout, resulting in a “bone against bone” joint, reduced motion and pain.Current therapeutic modalities are aimed at reducing pain and increasingjoint function. Non-invasive interventions such as exercise and weightloss are the first lines of treatment, followed by anti-inflammatorymedications. These latter treatments alleviate the symptoms but do notinhibit the processes that result in the changes characteristic of thisdisease and may actually accelerate joint destruction. Failure of thesetreatments usually culminates in surgical intervention (arthroplasty).Joint replacement is extremely successful with respect to restoringpatient mobility and decreasing pain. However, failure as a result ofosteolysis and aseptic loosening due to effects of wear debris orbiomechanically-related bone loss limit the lifetime of these implantsnecessitating higher-risk revision surgery at the expense of increasedpatient morbidity and failure rate. The present invention provides atreatment for osteoarthritis.

In traumatic rupture or detachment, the cartilage in the knee isfrequently damaged, and can be partially repaired through knee cartilagereplacement therapy.

In achondroplasia, reduced proliferation of chondrocytes in theepiphyseal plate of long bones during infancy and childhood results indwarfism.

Costochondritis is an inflammation of cartilage in the ribs, causingchest pain.

In spinal disc herniation, an asymmetrical compression of anintervertebral disc ruptures the sac-like disc, causing a herniation ofits soft content. The hernia often compresses the adjacent nerves andcauses back pain.

In relapsing polychondritis, a destruction, probably autoimmune, ofcartilage, especially of the nose and ears, causes disfiguration. Deathoccurs by suffocation as the larynx loses its rigidity and collapses.

Tumours made up of cartilage tissue, either benign or malignant, canoccur.

The present invention provides a treatment for each of the conditionsabove. Any of these conditions can be treated by repairing or growingnew cartilage or chondrocytes according to the methods disclosed hereinutilising a peptide comprising a C-terminus of a GH, or compositionthereof.

Other conditions that may be treated in accordance with the inventioninclude: chondromalacia patella; chondromalacia; chondrosarcoma-head andneck; chondrosarcoma; costochondritis; enchondroma; hallux rigidus; hiplabral tear; osteochondritis dissecans (OCD); osteochondrodysplasias;perichondritis; polychondritis; or torn meniscus.

The invention provides means to improve the function of existingchondrocytes and cartilage in maintaining a cartilaginous matrix. Italso provides means to promote growth of chondrocytes and cartilage andprovide a cartilaginous matrix, with or without an implant orprosthesis. In one embodiment the invention provides means to promotecartilage formation or repair in a cellular scaffold or in tissueengineering techniques, for example for cartilage generation or repairto grow new cartilage tissue in tissues including the nose, septum, ear,elbow, knee, ankle and invertebrate discs.

In one aspect the peptide is administered with an implant or the like toproduce or repair chondrocytes or cartilage tissue that may interactwith the implant to treat a condition as disclosed herein. As usedherein, “interact” refers to the effect in conjunction of components toachieve a desired biological outcome.

While not wishing to be bound by theory, when an implant “interacts”with chondrocytes, the effect of the implant in treating the conditionis greater than the effect of the implant alone and may be synergistic.

In one aspect the peptide is administered in combination withmesenchymal stem cells therapies to enhance repair. The effect oftreatment with the peptide and stem cells may be more than the additiveeffect of the separate treatments and may be synergistic.

In this embodiment, the “desired biological outcome” provided by theinvention is preferably cartilage repair and cartilage growth, morepreferably removal of the symptoms of osteoarthritis and most preferablytreatment and prevention of osteoarthritis.

In another example, the inventors show that a C-terminal growth hormonefragment, particularly AOD9604 (SEQ ID NO:1) can be used to promotemuscle growth, to improve recovery of muscle from injury, trauma or use,to improve muscle strength, to improve exercise tolerance, to increasethe proportion of muscle, to increase muscle mass, decrease musclewasting, improve muscle repair, or may be useful to treat disorders ofmuscle including wasting disorders, such as cachexia, and hormonaldeficiency, anorexia, AIDS wasting syndrome, sarcopenia, musculardystrophies, neuromuscular diseases, motor neuron diseases, diseases ofthe neuromuscular junction, and inflammatory myopathies in a subject inneed thereof.

The invention extends to treatment of disorders of muscle and ofdiseases associated with muscular degeneration characteristics. Nonlimiting examples of such disorders are various neuromuscular diseases,cardiac insufficiency, weakness of single muscles such as e.g. theconstrictor or bladder muscle, hypo- or hypertension caused by problemswith the constrictor function of vascular smooth muscle cells,impotence/erectile dysfunction, incontinence, AIDS-related muscularweakness, and general and age-related amyotrophia.

Disorders of muscle as referred to herein particularly include musclewasting conditions or disorders in which muscle wasting is one of theprimary symptoms.

“Muscle wasting” refers to the progressive loss of muscle mass and/or tothe progressive weakening and degeneration of muscles, including theskeletal or voluntary muscles which control movement, cardiac muscleswhich control the heart, and smooth muscles. In one embodiment, themuscle wasting condition or disorder is a chronic muscle wastingcondition or disorder. “Chronic muscle wasting” is defined herein as thechronic (i.e. persisting over a long period of time) progressive loss ofmuscle mass and/or to the chronic progressive weakening and degenerationof muscle. Chronic muscle wasting may occur as part of the agingprocess.

The loss of muscle mass that occurs during muscle wasting can becharacterized by a muscle protein breakdown or degradation, by muscleprotein catabolism. Protein catabolism occurs because of an unusuallyhigh rate of protein degradation, an unusually low rate of proteinsynthesis, or a combination of both. Protein catabolism or depletion,whether caused by a high degree of protein degradation or a low degreeof protein synthesis, leads to a decrease in muscle mass and to musclewasting. The term “catabolism” has its commonly known meaning in theart, specifically an energy burning form of metabolism.

Muscle wasting can occur as a result of age, a pathology, disease,condition or disorder. In one embodiment, the pathology, illness,disease or condition is chronic. In another embodiment, the pathology,illness, disease or condition is genetic. In another embodiment, thepathology, illness, disease or condition is neurological. In anotherembodiment, the pathology, illness, disease or condition is infectious.As described herein, the pathologies, diseases, conditions or disordersdirectly or indirectly produce a wasting (i.e. loss) of muscle mass,that is a muscle wasting disorder.

Also contemplated is the treatment of neuromuscular diseases which arealigned with joint or skeletal deformities. In one embodiment, musclewasting in a subject is a result of the subject having a musculardystrophy; muscle atrophy; or X-linked spinal-bulbar muscular atrophy(SBMA).

The muscular dystrophies are genetic diseases characterized byprogressive weakness and degeneration of the skeletal or voluntarymuscles that control movement. The muscles of the heart and some otherinvoluntary muscles are also affected in some forms of musculardystrophy. The major forms of muscular dystrophy (MD) are: Duchennemuscular dystrophy, myotonic dystrophy, Becker muscular dystrophy,limb-girdle muscular dystrophy, facioscapulohumeral muscular dystrophy,congenital muscular dystrophy, oculopharyngeal muscular dystrophy,distal muscular dystrophy and Emery-Dreifuss muscular dystrophy.

Muscular dystrophy can affect people of all ages. Although some formsfirst become apparent in infancy or childhood, others may not appearuntil middle age or later. Duchenne MD is the most common form,typically affecting children. Myotonic dystrophy is the most common ofthese diseases in adults.

Muscle atrophy (MA) is characterized by wasting away or diminution ofmuscle and a decrease in muscle mass. For example, Post-Polio MA is amuscle wasting that occurs as part of the post-polio syndrome (PPS). Theatrophy includes weakness, muscle fatigue, and pain.

Another type of MA is X-linked spinal-bulbar muscular atrophy (SBMA—alsoknown as Kennedy's Disease). This disease arises from a defect in theandrogen receptor gene on the X chromosome, affects only males, and itsonset is in adulthood.

Sarcopenia is a debilitating disease that afflicts the elderly andchronically ill patients and is characterized by loss of muscle mass andfunction. Further, increased lean body mass is associated with decreasedmorbidity and mortality for certain muscle-wasting disorders. Inaddition, other circumstances and conditions are linked to, and cancause muscle wasting disorders. For example, studies have shown that insevere cases of chronic lower back pain, there is paraspinal musclewasting.

Muscle wasting and other tissue wasting is also associated with advancedage. It is believed that general weakness in old age is due to musclewasting. As the body ages, an increasing proportion of skeletal muscleis replaced by fibrous tissue. The result is a significant reduction inmuscle power, performance and endurance.

Long term hospitalization due to illness or injury, or disusedeconditioning that occurs, for example, when a limb is immobilized, canalso lead to muscle wasting, or wasting of other tissue. Studies haveshown that in patients suffering injuries, chronic illnesses, burns,trauma or cancer, who are hospitalized for long periods of time, thereis a long-lasting unilateral muscle wasting, and a decrease in bodymass.

Injuries or damage to the central nervous system (CNS) are alsoassociated with muscle wasting and other wasting disorders. Injuries ordamage to the CNS can be, for example, caused by diseases, trauma orchemicals. Examples are central nerve injury or damage, peripheral nerveinjury or damage and spinal cord injury or damage. In one embodiment CNSdamage or injury comprise Alzheimer's diseases (AD); stroke, anger(mood); anorexia, anorexia nervosa, anorexia associated with agingand/or assertiveness (mood).

In another embodiment, muscle wasting or other tissue wasting (e.g.tendons or ligaments) may be a result of alcoholism.

In one embodiment, the wasting disease, disorder or condition beingtreated is associated with chronic illness

This embodiment is directed to treating, in some embodiments, anywasting disorder, which may be reflected in muscle wasting, weight loss,malnutrition, starvation, or any wasting or loss of functioning due to aloss of tissue mass.

In some embodiments, wasting diseases or disorders, such as cachexia,including cachexia caused by malnutrition, tuberculosis, leprosy,diabetes, renal disease, chronic obstructive pulmonary disease (COPD),cancer, end stage renal failure, emphysema, osteomalacia, orcardiomyopathy, may be treated by the methods of this invention

In some embodiments, wasting is due to infection with enterovirus,Epstein-Barr virus, herpes zoster, HIV, trypanosomes, influenza,coxsackie, rickettsia, trichinella, schistosoma or mycobacteria.

Cachexia is weakness and a loss of weight caused by a disease or as aside effect of illness. Cardiac cachexia, i.e. a muscle protein wastingof both the cardiac and skeletal muscle, is a characteristic ofcongestive heart failure. Cancer cachexia is a syndrome that occurs inpatients with solid tumours and haematological malignancies and ismanifested by weight loss with massive depletion of both adipose tissueand lean muscle mass.

Cachexia is also seen in COPD, acquired immunodeficiency syndrome(AIDS), human immunodeficiency virus (HIV)-associated myopathy and/ormuscle weakness/wasting is a relatively common clinical manifestation ofAIDS. Individuals with HIV-associated myopathy or muscle weakness orwasting typically experience significant weight loss, generalized orproximal muscle weakness, tenderness, and muscle atrophy.

Untreated muscle wasting disorders can have serious health consequences.The changes that occur during muscle wasting can lead to a weakenedphysical state resulting in poor performance of the body and detrimentalhealth effects.

Thus, muscle atrophy can seriously limit the rehabilitation of patientsafter immobilizations. Muscle wasting due to chronic diseases can leadto premature loss of mobility and increase the risk of disease-relatedmorbidity. Muscle wasting due to disuse is an especially serious problemin elderly, who may already suffer from age-related deficits in musclefunction and mass, leading to permanent disability and premature deathas well as increased bone fracture rate. Despite the clinical importanceof the condition few treatments exist to prevent or reverse thecondition. The inventors propose that the peptide can be used toprevent, repair and treat muscle wasting or atrophy associated with anyof the conditions recited above.

In a preferred embodiment the peptide is used to treat burns and sepsis.Human GH has been shown to have beneficial anabolic effects in treatingpatients after major surgery, trauma, sepsis, and burns.

Ramirez (1998) Ann Surg; 228, No. 4: 439-448 reports a study of thesafety and efficacy of hGH in the treatment of children who are severelyburned. The effect of hGH was manifested through elevated IGF-1 and thiscaused decreased glucose uptake, inhibition of glucose oxidation and adeficiency in glucose transport leading to elevated serum glucose. Whilethis can be treated with insulin this is not a desirable outcome. Theinventors propose that the methods of the present invention allowtreatment of burns without the associated side effects of elevated IGF-1as the C-terminal growth hormone fragment has the muscle anaboliceffects of human GH without containing an IGF-1 domain and thus withouthaving a measurable effect on IGF-1.

The invention in other aspects also contemplates treating healthyindividuals to cause an increase in muscle mass, strength, function oroverall physique. Full length growth hormone has been proposed topromote muscle recovery from injury or trauma or damage or overusethrough training and therefore to increase exercise tolerance. Theinventors propose that C-terminal growth hormone fragments may have thesame effect but without the side effects of elevated IGF-1.

The term “increase in muscle mass” refers to the presence of a greateramount of muscle after treatment relative to the amount of muscle masspresent before the treatment.

The term “increase in muscle strength” refers to the presence of amuscle with greater force generating capacity after treatment relativeto that present before the treatment.

The term “increase in muscle function” refers to the presence of musclewith greater variety of function after treatment relative to thatpresent before the treatment.

The term “increase in exercise tolerance” refers to the ability toexercise with less rest between exercise after treatment relative tothat needed before the treatment.

A muscle is a tissue of the body that primarily functions as a source ofpower. There are three types of muscles in the body: a) skeletalmuscle—striated muscle responsible for generating force that istransferred to the skeleton to enable movement, maintenance of postureand breathing; b) cardiac muscle—the heart muscle; and c) smoothmuscle—the muscle that is in the walls of arteries and bowel. Themethods of the invention are particularly applicable to skeletal musclebut may have some effect on cardiac and or smooth muscle. Reference toskeletal muscle as used herein also includes interactions between bone,muscle and tendons and includes muscle fibres and joints.

Peptides

The invention utilises a peptide comprising a carboxyl-terminal growthhormone fragment.

The terms carboxyl-terminal and C-terminal can be used interchangeably.

In one embodiment the invention utilises a peptide comprising acarboxyl-terminal fragment of human growth hormone. In one embodimentthe invention utilises a peptide comprising a carboxyl-terminal fragmentof a non-human growth hormone.

The published corresponding sequences of the C-terminal region of thegrowth hormone of selected mammals are tabulated below, using standardsingle letter notation. It will be apparent to persons skilled in theart that amino acid sequences of other animal growth hormones areavailable and they can also be used to design a peptide for use inaccordance with the invention.

GH Species Sequence SEQ ID human FRKDMDKVETFLR I VQCR SVEGSCGF  2human variant FRKDMDKVETFLR I VQCR SVEGSCGF  3 human CSFRKDMDKVETFLRMVQCR SVEGSCGF  4 monkey, rhesusFRKDMDK I ETFLR I VQCR SVEGSCGF  5 rat FKKDLHKAETYLRVMKCRRFAESSCAF  6mouse FKKDLHKAETYLRVMKCRRFAESSCAF  7 hamster FKKDLHKAETYLRVMKCRRFAESSCAF 8 whale, fin FKKDLHKAETYLRVMKCRRFVESSCAF  9 whale, seiFKKDLHKAETYLRVMKCRRFVESSCAF 10 fox, dog, cat FKKDLHKAETYLRVMKCRRFVESSCAF11 mink FKKDLHKAETYLRVMKCRRFVESSCAF 12 cattleFRKDLHKTETYLRVMKCRRFGEASCAF 13 sheep FRKDLHKTETYLRVMKCRRFGEASCAF 14 goatFRKDLHKTETYLRVMKCRRFGEASCAF 15 pig FKKDLHKAETYLRVMKCRRFVESSCAF 16 alpacaFKKDLHKAETYLRVMKCRRFVESSCAF 17 horse FKKDLHKAETYLRVMKCRRFVESSCAF 18elephant FKKDLHKAETYLRVMKCRRFVESSCAF 19 ancestral mammalFKKDLHKAETYLRVMKCRRFVESSCAF 20

The present invention also extends to the use of peptides which arefunctional homologues or variants of the native carboxyl-terminalsequences of human growth hormone or growth hormone of other animalspecies.

These functional homologues or variants may be derived by insertion,deletion or substitution of amino acids in, or chemical modification of,the native carboxyl-terminal sequence. Amino acid insertion variantsinclude amino and/or carboxylic terminal fusions as well asintra-sequence insertions of single or multiple amino acids. Insertionamino acid sequence variants are those in which one or more amino acidresidues are introduced into a predetermined site in the proteinalthough random insertion is also possible with suitable screening ofthe resulting product. Deletion variants are characterised by theremoval of one or more amino acids from the sequence. Substitution aminoacid variants are those in which at least one amino acid residue in thesequence has been replaced by another of the twenty primary proteinamino acids, or by a non-protein amino acid. In one embodimentsubstitutions are with conservative amino acids. Chemical modificationsof the native carboxyl-terminal sequence include the acetylation of theamino-terminus and/or amidation of the carboxyl-terminus and/or sidechain cyclisation of the native carboxyl-terminal sequence.

In one embodiment variants may comprise one, two, three, four or fiveinsertions, deletions or substitutions compared to the natural growthhormone C-terminal sequence provided that the function of the nativeC-terminal sequence is retained.

In one embodiment variants include a disulphide bond which confers acyclic configuration on the peptide. In particular, use of all of theactive peptides disclosed in AU 693478 and PCT/AU98/00724 is to beunderstood to be within the scope of this invention, for example:

Ref No. STRUCTURE 9502Leu Arg Ile Val Gln Pen Arg Ser Val Glu Gly Ser Pen Gly PheSEQ ID NO: 21 9405CH3CO-Leu Arg Ile Val Gln Cys Arg Ser Val Glu Gly Ser Cys Gly PheSEQ ID NO: 22 9410H-Leu Arg Ile Val Gln Cys Arg Ser Val Glu Gly Ser Cys Gly PheSEQ ID NO: 23 9404Leu Arg Ile Val Gln Cys Arg Ser Val Glu Gly Ser Cys Gly Phe-CONH₂SEQ ID NO: 24 9407Leu Arg Ile Val Gln Cys Lys Ser Val Glu Gly Ser Cys Gly PheSEQ ID NO: 25 9408Leu Arg Ile Val Gln Cys Lys Ser Val Glu Gly Ser Cys Gly PheSEQ ID NO: 26                           (amide bond) 9604Tyr Leu Arg Ile Val Gln Cys Arg Ser Val Glu Gly Ser Cys Gly PheSEQ ID NO: 1 9605Lys Leu Arg Ile Val Gln Cys Arg Ser Val Glu Gly Ser Cys Gly PheSEQ ID NO: 27 9618Lys Lys Leu Arg Ile Val Gln Cys Arg Ser Val Gln Gly Ser Cys Gly PheSEQ ID NO: 28 9607Ala Arg Ile Val Gln Cys Arg Ser Val G1u Gly Ser Cys Gly PheSEQ ID NO: 29 9606Leu Lys Ile Val Gln Cys Arg Ser Val Glu Gly Ser Cys Gly PheSEQ ID NO: 30 9608Leu Arg Ala Val Gln Cys Arg Ser Val Glu Gly Ser Cys Gly PheSEQ ID NO: 31 9403Leu Arg Lys Val Gln Cys Arg Ser Val Glu Gly Ser Cys Gly PheSEQ ID NO: 32 9609Leu Arg Ile Ala Gln Cys Arg Ser Val Glu Gly Ser Cys Gly PheSEQ ID NO: 33 9610Leu Arg Ile Val Ala Cys Arg Ser Val Glu Gly Ser Cys Gly PheSEQ ID NO: 34 9612Leu Arg Ile Val Gin Cys Arg Ala Val Glu Gly Ser Cys Gly PheSEQ ID NO: 35 9613Leu Arg Ile Val Gln Cys Arg Ser Ala Glu Gly Ser Cys Gly PheSEQ ID NO: 36 9615Leu Arg Ile Val Gln Cys Arg Ser Val Glu Ala Ser Cys Gly PheSEQ ID NO: 37 9616Leu Arg Ile Val Gln Cys Arg Ser Val Glu Gly Ala Cys Gly PheSEQ ID NO: 38 9602Leu Arg Ile Val Gln Cys Arg Ser Val Glu Gly Ser Cys Ala PheSEQ ID NO: 39 9501Leu Arg Ile Val Gln Cys Arg Ser Val Glu D-Ala Ser Cys D-Ala PheSEQ ID NO: 40 9601Leu Arg Ile Val Gln Cys Arg Ser Val Glu Gly Ser Cys Gly AlaSEQ ID NO: 41wherein the amino acid residue abbreviations used are in accordance withthe standard peptide nomenclature:

Gly=Glycine; Ile=Isoleucine; Glu=Glutamic Acid; Phe=Phenylalanine;Cys=Cysteine; Arg=Arginine; Gln=Glutamine; Leu=Leucine; Ser=Serine;Val=Valine;

Lys=Lysine; Ala=Alanine; Asp=Aspartic acid; His=Histidine;Orn=Ornithine;Tyr=Tyrosine; Pen=Penicillamine(p, p′-Dimethyl-Cysteine).

In one embodiment amino acids, except for glycine, are of the L-absoluteconfiguration. D configuration amino acids may also be used. The peptidemay have a cyclic disulfide bond between Cys(182) and Cys(189) orPen(182) and Pen(189) as appropriate.

Persons skilled in the art will appreciate that the peptide used may bemodified to improve storage stability, bioactivity, circulating halflife, or for any other purpose using methods available in the art, suchas glycosylation, by conjugation to a polymer to increase circulatinghalf-life, by pegylation or other chemical modification.

For example it may be desirable to introduce modification to improvestorage stability or to improve bioavailability.

In one embodiment the peptide comprises amino acids 182-189 (hGH182-189), amino acids 181-189, 180-189, 179-189, 178-189, 177-189,176-189, 175-189, 175-190, 176-190, 177-190, 178-190, 179-190, 180-190,181-190, 182-190, 182-191, 181-191, 180-191, 179-191, 178-191, 177-191,176-191 or 175-191 of human growth hormone or corresponding peptidesfrom growth hormone of non-human animals, for example from within SEQ IDNO: 2-20.

In one embodiment the peptide comprises a variant of a C-terminalfragment of hGH comprising amino acids 182-189 or particularly 177-191.In a preferred embodiment the peptide is AOD9604 (Tyr-hGH 177-191) SEQID NO: 1.

“Peptide” as used herein means any chain of amino acids from 8 to 50amino acid residues in length, preferably 8 to 40, 8 to 30, 8 to 25, or8 to 20, or more preferably about 8, 9, 10, 11, 12, 13, 14, 15, 16, 17,18 or 19 amino acid residues in length.

Any peptide for use in accordance with the present invention cannot havethe full length sequence of a growth hormone. Full length growth hormoneis effective for the treatments proposed but also increases secretion ofIGF-1. Accordingly full length growth hormone does not fall within thescope of the peptides for use in accordance with the present invention.

Any peptide for use in accordance with the present invention cannot havethe full length sequence of a growth hormone as the peptide for use inthe present invention cannot contain the domain of growth hormoneresponsible for IGF-1 production. As used herein the domain of growthhormone responsible for IGF-1 production is amino acid residues 6-14 ofhuman growth hormone.

A peptide for use in accordance with the present invention may bederived from natural sources, produced by recombinant DNA technology, orsynthesised using conventional peptide synthetic methods.

The peptide may be conjugated to a fusion partner to enable easierbiosynthesis and/or delivery. It may be incorporated in a conventionalpharmaceutical composition, or may be present in a genetically-modifiedfood, such as disclosed in WO 01/33997.

Administration

The peptide may be administered in a pharmaceutical composition togetherwith a pharmaceutically acceptable carrier for administration.

The peptide may be administered by any suitable route, and the personskilled in the art will readily be able to determine the most suitableroute and dose for the condition to be treated and the subject. Thepeptide may be administered orally, sublingually, buccally,intranasally, by inhalation, transdermally, topically, intra-articularlyor parenterally in dosage unit formulations containing conventionalnon-toxic pharmaceutically acceptable carriers, adjuvants, and vehicles.The term parenteral as used herein includes subcutaneous, intravenous,intramuscular, intrathecal, intracranial, injection or infusiontechniques.

In one embodiment the peptide may be administered with or in an implant,medical device or prosthesis. The implant may be a biodegradable implantor slow release depot or other implant as known to persons skilled inthe art. Such embodiment is particularly appropriate for improvingmuscle growth and strength after muscle trauma or damage.

When used to treat burns the peptide may be administered orally,topically or parenterally.

Compositions comprising the peptide are to be administered in atherapeutically effective amount. As used herein, an “effective amount”is a dosage which is sufficient to reduce to achieve a desiredbiological outcome. The desired biological outcome may be anytherapeutic benefit including an increase in muscle mass, an increase inmuscle strength, muscle growth, or treatment of burns or wounds. Suchimprovements may be measured by a variety of methods including thosethat measure lean and fat body mass (such as duel ray scanninganalysis), muscle strength, or the formation of muscle cells.

A typical daily dosage might range from about 1 μg/kg to up to 100 mg/kgor more, depending on the mode of delivery.

Dosage levels of the growth hormone fragment could be of the order ofabout 0.1 mg per day to about 50 mg per day, or will usually be betweenabout 0.25 mg to about 1 mg per day. The amount of active ingredientwhich may be combined with the carrier materials to produce a singledosage will vary, depending upon the host to be treated and theparticular mode of administration. For example, a formulation intendedfor oral administration to humans may contain about 1 mg to 1 g of anactive compound with an appropriate and convenient amount of carriermaterial, which may vary from about 5 to 95 percent of the totalcomposition. Dosage unit forms will generally contain between from about0.1 mg to 50 mg of active ingredient.

It will be understood, however, that the specific dose level for anyparticular subject will depend upon a variety of factors including theactivity of the specific compound employed, the age, body weight,general health, sex, diet, time of administration, route ofadministration, rate of excretion, drug combination and the severity ofthe particular disease undergoing therapy.

Dosage schedules can be adjusted depending on the half life of thepeptide, or the severity of the subject's condition.

Generally, the compositions are administered as a bolus dose, tomaximize the circulating levels of peptide for the greatest length oftime after the dose. Continuous infusion may also be used after thebolus dose.

Subject

The treatments of the present invention are suitable for subjects inneed thereof. “Subject,” as used herein, refers to human and non-humananimals.

The term “non-human animals” includes all vertebrates, e.g., mammals,such as non-human primates (particularly higher primates), sheep, horse,dog, rodent (e.g., mouse or rat), guinea pig, goat, pig, cat, rabbits,cow, and non-mammals, such as chickens, amphibians, reptiles, etc. Inone embodiment, the subject is an experimental animal, an animalsuitable as a disease model, or in animal husbandry (animals as foodsource), where methods to increase lean muscle mass will greatly benefitthe industry. Additionally the method of the first aspect isparticularly important in race horses.

In one embodiment the treatment is for humans, particularly adulthumans, children aged 11 to 16 years old, aged 4 to 10 years old,infants of 18 months up to 4 years old, babies up to 18 months old. Thetreatment may also be used for elderly or infirm humans.

In one embodiment the subject is suffering from a disease, for exampleosteoarthritis, diabetes, HIV, or is immunocompromised. The subject maypossess pre-diabetic markers or have a disease in which diabetes in acomplication. In another embodiment the subject is desirous of animprovement in their health or appearance, for example to slow aging orimprove physique. Subjects therefore include athletes, both elite andamateur, body builders, those desirous of enhanced physique, combatantsand manual workers.

In one embodiment the treatments of the present invention are used tosupplement alternative treatments for the same condition. For examplethe C-terminal growth hormone fragment can be used to supplement stemcell therapies for joint and muscle repair.

In one embodiment the subject is contraindicated for growth hormonetherapy.

Throughout this specification and claims which follow, unless thecontext requires otherwise, the word “comprise”, or variations such as“comprises” or “comprising”, will be understood to imply the inclusionof a stated integer or group of integers but not the exclusion of anyother integer or group of integers.

It must also be noted that, as used in the subject specification, thesingular forms “a”, “an” and “the” include plural aspects unless thecontext clearly dictates otherwise.

It will be apparent to the person skilled in the art that while theinvention has been described in some detail for the purposes of clarityand understanding, various modifications and alterations to theembodiments and methods described herein may be made without departingfrom the scope of the inventive concept disclosed in this specification.

The invention will now be described by way of reference only to thefollowing non-limiting examples.

EXAMPLES Example 1: Determine the Effect of AOD on Cartilage Repair byEvaluating Tissue Formation Methods Cell Culture and Treatment

Chondrocytes were isolated from articular cartilage obtained asepticallyfrom calf metacarpal-carpal joints and seeded onto collagen typeII-coated membrane inserts (Millipore™) (1.5×10⁶ cells/12 mm diametermembrane). Chondrocytes were grown in Ham's F-12 supplemented with 5%fetal bovine serum (FBS) under standard culture conditions. After 5 daysthe serum was increased to 20%. Chondrocytes were then grown in thepresence and absence of AOD9604 (SEQ ID NO: 1) (10 or 100 μg/ml) for 2weeks (total culture time 3 weeks). Fresh AOD was added with each changeof media.

Assessment of Tissue

The tissue was harvested and digested with papain (Sigma, 40 μg/ml) for48 h at 65° C. DNA content was determined using the Hoechst 33258 dyebinding assay (Polysciences) and fluorometry (excitation 365 nm,emission 458 nm). Proteoglycan content was determined by measuring theamount of sulphated glycosaminoglycans using the dimethylmethylene bluedye binding assay and spectrophotometry (525 nm). Collagen content wasdetermined by measuring the hydroxyproline content of an aliquot of thepapain digest hydrolyzed in 6N HCl for 18 h at 110° C. Thehydroxyproline content was determined using the chloramine-T/Ehrlich'sreagent assay and spectrophotometry (560 nm). The cellularity wasquantified by measuring the DNA content using Hoescht 33258 dye bindingassay (Polysciences) and fluorometry (excitation 365 nm, emission 458nm).

Statistical Analysis

All experiments were done three times and each condition was done intriplicate. The data was expressed as mean±SD. The data was expressed asmean±SD. Significance was determined using two way ANOVA followed byTukey's post hoc test when multiple groups were being evaluated. T testwas utilized if 2 groups were being compared. Significance was assignedat p values <0.05.

Results

AOD9604 treatment was not toxic to the cells as determined byquantifying DNA content after 2 weeks of treatment (FIG. 1).

AOD9604 (100 μg/ml) enhanced cartilage repair by increasing proteoglycanand collagen content of the tissue as determined at 3 weeks of culture(FIG. 2 and FIG. 3).

In the figures AOD9604 is referred to as AOD.

Example 2: Effect of AOD on Native Cartilage Methods

Cartilage explants were taken from calf metacarpal-carpal joints andplaced in culture in Ham's F-12 supplemented with 20% fetal bovine serum(FBS) in the presence and absence of AOD9604 (SEQ ID NO: 1) (10 or 100μg/ml) for 1 week or (100 or 500 μg/ml) for 2 weeks.

Assessment of Tissue

The tissue was harvested and water content and dry weight measured. Thetissue was then digested with papain (Sigma, 40 μg/ml) for 48 h at 65°C. DNA content was determined using the Hoechst 33258 dye binding assay(Polysciences) and fluorometry (excitation 365 nm, emission 458 nm).Proteoglycan content was determined by measuring the amount of sulphatedglycosaminoglycans using the dimethylmethylene blue dye binding assayand spectrophotometry (525 nm). Collagen content was determined bymeasuring the hydroxyproline content of an aliquot of the papain digesthydrolyzed in 6N HCl for 18 h at 110° C. The hydroxyproline content wasdetermined using the chloramine-T/Ehrlich's reagent assay andspectrophotometry (560 nm). The cellularity was quantified by measuringthe DNA content using Hoescht 33258 dye binding assay (Polysciences) andfluorometry (excitation 365 nm, emission 458 nm.

Statistical Analysis

All experiments were done three times and each condition was done intriplicate. The data was expressed as mean±SD. The data was expressed asmean±SD. Significance was determined using two way ANOVA followed byTukey's post hoc test when multiple groups were being evaluated. T testwas utilized if 2 groups were being compared. Significance was assignedat p values <0.05.

Results

AOD was not toxic to cells at concentrations up to 500 μg/ml for 2 weeksof treatment as there was no significant change in DNA content in thattime period (FIG. 4).

AOD9604 had no effect on native cartilage at concentrations of AOD of500 μg/ml (FIGS. 5-8).

In the figures AOD9604 is referred to as AOD.

Conclusions

1) AOD9604 had a positive (anabolic) on cartilage tissue formationsuggesting AOD would be appropriate to use to stimulate cartilagerepair.

2) AOD9604 has no effect on intact cartilage.

Example 3: Effect of Peptide AOD9604 on Myoblast Differentiation intoMuscle Cells In Vitro Methods Cell Culture and Treatment

C2C12 were grown in monolayer culture in DMEM (high glucose)supplemented with 10% fetal bovine serum under standard cell cultureconditions. The cells were passaged at 60-70% confluence. The effect ofAOD9604 (SEQ ID NO: 1) on cell proliferation was determined by growingthe cells (3000 cells/cm²) in the absence or presence of AOD9604 (10 and100 μg/ml) for 3 days. The DNA content was then quantified. The cellswere digested by papain and DNA content determined using the Hoechst33258 dye binding assay (Polysciences) and fluorometry (excitation 365nm, emission 458 nm).

Assessment of Cell Differentiation

C2C12 (2×10³ cells/cm²) were grown in DMEM and 10% FBS. Differentiationwas induced by replacing growth media with differentiation media whichconsists of DMEM supplemented with 2% horse serum. The cells were grownin the absence or presence of AOD9604 (10 or 100 μg/ml). The cells werefixed in 4% paraformaldehyde and co-stained for myogenin (a musclespecific muscle marker (transcription factor) 1/250 Tris-Triton Xbuffer) and Bodipy (cytoplasmic stain, Invitrogen, 1/1000 PBS). Thenumber of cells stained for myogenin were counted in 4 random fields (upto 100 cells/field) (n=4). The percent of cells that stained positivelyfor myogenin was calculated.

For myotube quantification the cells were stained with Bodipy(Invitrogen, 1/1000 PBS) according to the manufacturer's directions. Thenumber of myotubes that developed over 72 hours was determined andexpressed as a percent of total cells in culture (n=3, 100cells/culture).

Statistical Analysis

The data was expressed as mean±SD. The data was expressed as mean±SD.Significance was determined using two way ANOVA followed by Tukey's posthoc test when multiple groups were being evaluated. T test was utilizedif 2 groups were being compared. Significance was assigned at p values<0.05.

Results

AOD9604 enhances C2C12 differentiation into myoblasts by 72 hours atdoses as low as 10 μg/ml (FIGS. 9 and 10). AOD9604 had no affect on theformation of myotubes (FIG. 11). AOD9604 did not affect cellproliferation (FIG. 12). In the figures AOD9604 is referred to as AOD.

Conclusion

AOD9604 enhances differentiation to myoblasts and may be appropriate touse to enhance muscle formation.

Example 4: Effect of Intra-Articular Administration of AOD9604 in aRabbit Collagenase Model of Osteoarthritis Given Alone and inCombination with HA Study Design

The study followed the protocols as described in Kim et al., (2010) JKorean Med Sci 25:776-780, which demonstrated the therapeutic effect ofintra-articular administration of hyaluronic acid (HA) with and withoutAOD9604 (SEQ ID NO:1). AOD9604 (SEQ ID NO: 1) was provided by MetabolicPharmaceutical Pty Ltd.

Species: New Zealand White Rabbit Sex: Male

Age: 12 weeks

Number of Rabbits: 32

Treatments groups: 32 rabbits were each given a 2 mg collagenaseinjection into their right knee joint, followed by a second injection 3days later to induce osteoarthritis. After 4 weeks, the rabbits weredivided into 4 groups of 8 for subsequent treatments.

Group 1 received 4 weekly injections of vehicle (disease control group).

Group 2 received 4 weekly injections of 6 mg of HA-hyuran-plus (HApositive control group).

Group 3 received 4 weekly injections of 0.25 mg of AOD9604 (AOD onlygroup).

Group 4 received 4 weekly injections of both HA and AOD9604 (combinationgroup).

Clinical observations were made by direct observation daily to assesslameness (as described in Kim et al., (2010) supra). At 9 weeks postinitial collagenase injection the animals were euthanized.

Gross observation of the right and left knee joints was performed afterdissection and scored for damage (as described in Kim et al., (2010)supra). Histological analysis of lateral and medial condyles from eachjoint was performed and scored.

A statistical analysis of the data was performed and the results areshown in FIGS. 13 (Group 1), 14 (Group 2), 15 (Group 3) and 16 (Group4). The results indicate that AOD9604 (SEQ ID NO:1) has regenerativepower compared to control and that HA and AOD9694 (SEQ ID NO: 1) providea similar amount of improvement at this point in time.

Example 5: Case Study—Infection

The subject was a 46 year old male. Nail puncture in the left foot. Thesubject described a very swollen foot with a high degree of redness fortwo centimeter circumference around the punctured site. There was also ahigh degree of tenderness. The swelling was identified throughout theright side of the left foot and through the fourth and fifth phalanges.Some swelling was noted back into the ankle joint. Symptoms consistentwith Clostridium tetani infection.

Dosage:

Three 360 mcg doses of AOD9604 (SEQ ID NO:1) subcutaneously around theinfected area per day. Each individual dose represented an intake of 0.3ml at 1200 mcg per ml per dose. 4 days.

Main Observations:

By day five the subject was very happy with both the pain andfunctionality in the region. He was able to fully cloth the foot with ashoe and could resume general weight bearing activity. The subjectresumed full running and moderate weight lifting activity on day eight.The subject also noted that the pain was considerably reduced by aroundthree of the injections. The majority of this pain reduction wasassociated with the swelling from the infection. The tissue discomfortdue to disruption resolved at around day five. The subject felt allsymptoms and any other associated minor discomfort had fully resolved ataround day eight. No other chemotherapeutic intervention orantimicrobiotics used in the treatment.

Example 6: Case Study—Calf Muscle

The subject was a 24 year old male professional footballer. The athleteincurred at 1.5 cm tear on his soleus near the adjacent aponeurosis tothe gastrocnemius. The MRI displayed some associated bleeding andcontusion. A 1.5 cm tear is considered a moderately large tear for theregion. The MRI indicated that a mass of blood and edema had positioneditself near the anterior aponeurosis insertion onto the median septum.The fibre tears were directional with the orientation of the fibres. Thelength of the tear was 1.5 cm.

Dosage:

The subject undertook an intervention of AOD9604 (SEQ ID NO: 1) at 600mcg per dose. He applied one ml twice daily. The transdermal applicationwas at 600 mcg per ml. The total application was 20 ml.

Main Observations:

The athlete's injury was resolved in two weeks. This was considerably areduction in time frame given the original nature of the injury. The MRIconfirmed that that full integrity had returned to the injury. Nobleeding or edema was identified on the follow up MRI. Also at thispoint the athlete had returned to full weight bearing activities. Theathlete indicated that there was no residual pain or discomfortassociated with the original injury. There was no awareness of the areawith the return to exercise. There was no immediate pain or discomfortonce the weight bearing capacity was fully resumed. The athlete alsonoted that the pain associated with the injury had started to diminishquite rapidly after the injury application of the transdermal. Heindicated that there was no resumption of any pathology and in relationto pain or inflammation on return. He was comfortable with runningmechanics and he commented that no discomfort was noted. The athleteconsidered that the injury had returned to full function.

Example 7: Case Study—Hamstring Muscle

The subject was a 22 year old male professional footballer. The athleteincurred at 2.0 cm tear to the long head of his bicep femoris. The MRIindicated that a mass of blood and edema was prominant near the upperthird of the long head muscle belly. Some fibre disruption was notedbelow the tendon junction coming from the insertion. The length of thetear was 1.5 cm. The fusiform belly region around the injury was highlyinflamed.

Dosage:

The subject undertook an intervention of AOD9604 (SEQ ID NO: 1) at 600mcg per dose. He applied one ml twice daily. The transdermal applicationwas at 600 mcg per ml. The total application was 40 ml.

Main Observations.

The athlete's injury was resolved in three weeks. It was considered thatwas a significant reduction in the time frame expected given theoriginal pathology of the injury. The MRI confirmed that that fullintegrity had returned to the bicep femoris long head. No residualbleeding or edema was identified on the follow up MRI. The athlete hadreturned to full weight bearing activities at the conclusion of thethree week application of the transdermal. This was thought to be aheadof schedule. The athlete noted that there was no residual pain ordiscomfort associated with the original injury once returning to fullweight bearing duties. There was no awareness of the area with thereturn. There was no immediate pain or discomfort identified with thefull weight bearing capacity. The athlete felt that the pain associatedwith the injury had started to diminish within the first week ofapplication of the transdermal. He indicated that there was noaggravation of pathology and no pain or inflammation. He had fullcomfort with the running mechanics and no discomfort was noted during orafter exercise. Within the first week of the return to full weightbearing activities the athlete experienced no limitations to weightbearing capacity and was considered to have a full return to function.

Example 8—Case Study—Shoulder Tendon

The subject was a 23 year old male professional footballer. The athleteincurred some fibre disruption at the junction of supraspinatus tendoninsertion. The MRI indicated an area of 1.0 cm within the disruptivepathology. The MRI indicated that the disruption approached mostsuperior facet of the greater tubercle of the humerus. Most of theaffected area was the muscular portion of the junction. The MRIindicated an area of 1.0 cm within the disruptive pathology. The athleteindicated that the pain associated with the area was noticeable foraround 5 weeks indicating some chronicity to the injury. The athlete hadbeen restricted in both the range of motion he could employ for theinjured site as well as the amount of weight bearing activity he couldengage for the area. He had been finding it difficult to abduct the armat the shoulder joint. He found difficulty in resisting inferiorgravitational forces placed across the shoulder joint and foundaggravation with any downward pull with weight of the upper limb.

Dosage:

The subject undertook an intervention of AOD9604 (SEQ ID NO: 1) at 600mcg per dose. He applied one ml twice daily. The transdermal applicationwas at 600 mcg per ml. The total application was 30 ml.

Main Observations:

The athlete's injury was resolved in three weeks. Subsequent MRIconfirmed that that full integrity had returned to the junctional area.The athlete also had returned to full weight bearing activities androtation activity with the area. The athlete felt that there was noresidual pain or discomfort associated with either movement or resistantwork with the region. There was no awareness of the area with the returnto exercise in either the rotation or weight bearing components. Theathlete confirmed that there was no immediate pain or discomfort oncethe weight bearing capacity was fully resumed and no pain, inflammationor awareness of the injury could be identified post training. There wasre-aggravation of the injury at any stage. The athlete also noted thatthe pain associated with the injury had started to diminish by the endof the first week of the application of the transdermal.

Example 9—Case Study—Corked Quad—Hematoma

The subject was a 24 year old male professional footballer. The athleteincurred a contusion to the rectus femoris adjacent to the centraltendon. The MRI indicated an area of 2.0 cm area for the contusion. TheMRI indicated that there was fibre disruption within the middle of therectus femoris just adjacent to the region of the central tendon. TheMRI indicated that the area of disruption was 2.0 cm. There was a minordegree of associated edema associated with the pathology. The MRIconfirmed that there was fibre disruption but no true tear. The centraltendon remained intact. The injury was an acute injury. The athleteidentified discomfort on weight bearing movement and the mass was easilypalpatable. Any hip flexion movement could induce discomfort andawareness of the injury. Although the muscle is not considered adominant muscle in knee extension movement the affected area was painfuland inducing discomfort with such work.

Dosage:

The subject undertook an intervention of AOD9604 (SEQ ID NO: 1) at 600mcg per dose. He applied one ml twice daily. The transdermal applicationwas at 600 mcg per ml. The total application was 20 ml.

Main Observations:

The athlete's injury was resolved in two weeks. The area for thecontusion was noted to be clear of any bleeding or edema on subsequentMRI. The MRI confirmed a return to full anatomical integrity for theinjured area. The athlete returned to full weight bearing activityexperiencing no issues with mechanical load or running mechanics. Therewere no issues with resumption of contact. The athlete confirmed thatthere was no immediate pain or discomfort once the weight bearingcapacity was fully resumed, The athlete felt that no pain, inflammationor awareness of the injury could be identified post training. There wasno re-aggravation of the injury at any stage either through running,weight bearing or contact. The athlete noted that the pain associatedwith the injury had started to diminish by the end of the first week ofthe application of the transdermal though some mass could still palpatedin the injured area.

1-11. (canceled)
 12. A method for improving muscle, ligament or tendonmass, repair, form or function in a subject, the method comprisingadministering to a subject in need of such treatment an effective amountof a peptide that is up to 50 amino acid residues in length andcomprises amino acid residues 182-189 of human growth hormone or thecorresponding region from any one of SEQ ID Nos: 1-41, which peptidedoes not include the domain of growth hormone responsible for IGF-1production.
 13. The method of claim 20, wherein the peptide comprises acarboxyl-terminal sequence from human growth hormone or a carboxylterminal sequence from a growth hormone of a non-human animal.
 14. Themethod of claim 20, wherein the peptide is administered in combinationwith mesenchymal stem cell therapy.
 15. The method of claim 20, whereinthe peptide is administered to supplement stem cell therapy for jointand muscle repair.
 16. The method of claim 20, wherein the peptidecomprises amino acids 182-189 of human growth hormone.
 17. The method ofclaim 20, wherein the peptide comprises amino acids 177-191 of humangrowth hormone.
 18. The method of claim 20, wherein the peptidecomprises a sequence selected from SEQ ID NO: 1-5, 22-24, 27-34 39 and41.
 19. The method of claim 20, wherein the peptide consists of asequence selected from SEQ ID NO: 1-5, 22-24, 27-34 39 and
 41. 20. Themethod of claim 20, wherein the peptide is administered byintra-articular administration with hyaluronic acid.